Abstract

Introduction Effective cancer therapy is hampered by the development of multidrug resistance by most cancer types. Cancer Stem Cells (CSCs) which are Side Population (SP) cells from tumors are attributed to the drug resistance phenotype. Efficient therapies for the successful eradication of tumor would have to employ agent(s) capable of inhibiting the self-renewal pathways and blocking or avoiding the drug-efflux property of these CSCs. Experimental Procedures Cell culture: The H1650 mixed population cells were maintained in DMEM:F12 (50:50) containing 2% PSN and supplemented with 10% fetal bovine serum (FBS) in the presence of 5% CO2 at 37[[Unsupported Character - &#8304;]]C. The H1650 SP cells were cultured in DMEM:F12 (50:50) containing, 1x Nitrogen supplement, 10 µg/mL each of bFGF and EGF. In vitro cytotoxicity studies: The crystal violet dye assay was used to determine the viability of both cell types 72 hours after treatment with test compounds. Cell invasion assay: The invasion of cells through a basement membrane matrix was assessed using the Cultrex 96-well collagen I cell invasion assay from Trevigen according to manufacturer's protocol. Cell migration assay: Migration of cells was determined using the wound healing assay before and after 24 hours of incubation post-treatment. Results The H1650 mixed population cells produced IC50 values of 11.03 µM, 2.68 µM, 8.70 µM, 2.63 µM, 6.48 µM, 5.06 µM, and 6.82 µM after treatment with Cisplatin, Gemcitabine, Doxorubicin, Fluorouracil, Camptothecin, DIM-5 and DIM-8, respectively. Significantly higher IC50 values were observed after similar treatments with Cisplatin (142.01 µM), Gemcitabine (42.68 µM), Doxorubicin (133.76 µM), Fluorouracil (81.01 µM), Camptothecin (48.51 µM), DIM-5 (15.67 µM) and DIM-8(19.69 µM). The invasiveness of the side population cells was seen to be generally diminished following treatment with 25 µM of test compound with percentage invasion ranging between 13.07% and 66.63% and a mean invasion of 37.92%. Further, the side population cells exhibited a much greater migratory index relative to the mixed population cells. Conclusions The resistance of the H1650 stem cells to chemotherapy is evident from the comparatively high IC50 values.; the exception being DIM-5 and DIM-8 exhibiting about 2 to 7 times more anticancer effect. These two agents act on the TR3/Nur77 nuclear receptor as activators and deactivators of the receptor, respectively, with resultant apoptotic events occurring downstream. With such demonstrable anticancer activities, the DIM compound may be potentially useful in sensitizing H1650 stem cells to chemotherapy when given in combination.